LOCUS       sequence-472892-        7623 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Sleeping Beauty transposon plasmid for hydrodynamic tail vein 
            injection (HDTVi) into mice. Caggs promotor driven expression of 
            mVenus and mutant NRAS (NRASG12V)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7623)
  AUTHORS   Chan ASL, Zhu H, Narita M, Cassidy LD, Young ARJ, Bermejo-Rodriguez 
            C, Janowska AT, Chen HC, Gough S, Oshimori N, Zender L, Aitken SJ, 
            Hoare M, Narita M
  TITLE     Titration of RAS alters senescent state and influences tumour 
            initiation.
  JOURNAL   Nature. 2024 Sep;633(8030):678-685. doi: 10.1038/s41586-024-07797-z.
            Epub 2024 Aug 7.
  PUBMED    39112713
REFERENCE   2  (bases 1 to 7623)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41586-024-07797-z"; journalName: "Nature"; date: 
            "2024-09"; volume: "633"; issue: "8030"; pages: "678-685"
COMMENT     Sequence Label: pT/CAGGS-NRASG12V-IRES-mVenus
FEATURES             Location/Qualifiers
     source          1..7623
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             36..53
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     misc_feature    658..1210
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(802..819)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1029..1048
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(1222..1224,1225..1227,1228..1941)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1267..1288)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1528..1547)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1875..1896
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2123..2140)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2129..2353
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2800..2816)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2800..2816)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2813..2835)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2824..2840
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2848..2854,2855..2872,2873..2878))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2893..2914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3031..3048)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3202..3790)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3282..3301)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3961..4752,4753..4821))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4584..4603
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4822..4926)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4994..5012
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5050..5072)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5172..5191
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5385..5407
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5399..5416
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5400..5416
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5879..5895)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5880..5896)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     enhancer        5895..6274
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6276..6551
                     /label=chicken beta-actin promoter
     intron          6552..7568
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     7492..7514
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     7576..7595
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
ORIGIN
        1 tcgataccgg gggcgcgtcc accatggagg aagaggaata tatgcctatg gaaatgactg
       61 agtacaaact ggtggtggtt ggagcagttg gtgttgggaa aagcgcactg acaatccagc
      121 taatccagaa ccactttgta gatgaatatg atcccaccat agaggattct tacagaaaac
      181 aagtggttat agatggtgaa acctgtttgt tggacatact ggatacagct ggacaagaag
      241 agtacagtgc catgagagac caatacatga ggacaggcga aggcttcctc tgtgtatttg
      301 ccatcaataa tagcaagtca tttgcggata ttaacctcta cagggaacag attaagcgag
      361 taaaagactc ggatgatgta cctatggtgc tagtgggaaa caagtgtgat ttgccaacaa
      421 ggacagttga tacaaaacaa gcccacgaac tggccaagag ttacgggatt ccattcattg
      481 aaacctcagc caagaccaga cagggtgttg aagatgcttt ttacacactg gtaagagaaa
      541 tacgccagta ccgaatgaaa aaactcaaca gcagtgatga tgggactcag ggttgtatgg
      601 gattgccatg tgtggtgatg taagcggccg accmggtmcc cccccccccc ccccctaacg
      661 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca
      721 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga
      781 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga
      841 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca
      901 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag
      961 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa
     1021 gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc cagaaggtac
     1081 cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt gtttagtcga
     1141 ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac
     1201 gatgataata tggccacaac catggtgagc aagggcgagg agctgttcac cggggtggtg
     1261 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     1321 ggcgagggcg atgccaccta cggcaagctg accctgaagc tgatctgcac caccggcaag
     1381 ctgcccgtgc cctggcccac cctcgtgacc accctgggct acggcctgca gtgcttcgcc
     1441 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1501 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     1561 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1621 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     1681 accgccgaca agcagaagaa cggcatcaag gccaacttca agatccgcca caacatcgag
     1741 gacggcggcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1801 gtgctgctgc ccgacaacca ctacctgagc taccagtcca agctgagcaa agaccccaac
     1861 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1921 atggacgagc tgtacaagta agtcgacctg cagccaagct tatcgataaa ataaaagatt
     1981 ttatttagtc tccagaaaaa ggggggaatg aaagacccca cctgtaggtt tggcaagcta
     2041 gcattcccgg gaaaactcaa cagcagtgat gatgggactc agggttgtat gggattgcca
     2101 tgtgtggtga tgtaaggatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2161 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2221 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2281 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     2341 gtgggctcta tggcttctga ggcggaaaga accagccatc acaaagctct gacctcaatc
     2401 ctatagaaag gaggaatgag ccaaaattca cccaacttat tgtgggaagc ttgtggaagg
     2461 ctactcgaaa tgtttgaccc aagttaaaca atttaaaggc aatgctacca aatactaatt
     2521 gagtgtatgt taacttctga cccactggga atgtgatgaa agaaataaaa gctgaaatga
     2581 atcattctct ctactattat tctgatattt cacattctta aaataaagtg gtgatcctaa
     2641 ctgaccttaa gacagggaat ctttactcgg attaaatgtc aggaattgtg aaaaagtgag
     2701 tttaaatgta tttggctaag gtgtatgtaa acttccgact tcaactgtag gggatcctct
     2761 agagtcgacc tgcaggcatg caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     2821 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     2881 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     2941 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     3001 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     3061 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     3121 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     3181 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     3241 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     3301 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     3361 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     3421 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     3481 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     3541 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     3601 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     3661 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     3721 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3781 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3841 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3901 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3961 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     4021 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     4081 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4141 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     4201 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     4261 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     4321 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     4381 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     4441 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     4501 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     4561 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     4621 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     4681 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     4741 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4801 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4861 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4921 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     4981 attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga
     5041 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     5101 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     5161 gcttaactat gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat
     5221 accgcacaga tgcgtaagga gaaaataccg catcaggcgc cattcgccat tcaggctgcg
     5281 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
     5341 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
     5401 taaaacgacg gccagtgaat tcgagctcgg taccctacag ttgaagtcgg aagtttacat
     5461 acacttaagt tggagtcatt aaaactcgtt tttcaactac tccacaaatt tcttgttaac
     5521 aaacaatagt tttggcaagt cagttaggac atctactttg tgcatgacac aagtcatttt
     5581 tccaacaatt gtttacagac agattatttc acttataatt cactgtatca caattccagt
     5641 gggtcagaag tttacataca ctaagttgac tgtgccttta aacagcttgg aaaattccag
     5701 aaaatgatgt catggcttta gaagcttctg atagactaat tgacatcatt tgagtcaatt
     5761 ggaggtgtac ctgtggatgt atttcaaggc gatgtacggg ccagatatac gcgttgacat
     5821 tgattattga ctagtaacgg ccgccagtgt gctggaattc tgcagatatc aagcttatcg
     5881 ataccgtcga cctcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
     5941 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
     6001 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
     6061 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     6121 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     6181 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
     6241 gtacatctac gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt
     6301 cactctcccc atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt
     6361 attttgtgca gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg
     6421 gcgaggggcg gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc
     6481 tccgaaagtt tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg
     6541 cgcggcgggc gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct
     6601 cgcgccgccc gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg
     6661 gcccttctcc tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt
     6721 ggctgcgtga aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg
     6781 ggggtgcgtg cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg
     6841 gctgtgagcg ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga
     6901 gcgcggccgg gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg
     6961 tgcggggtgt gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac
     7021 cccccctgca cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc
     7081 gtacggggcg tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc
     7141 cgggcggggc ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccc
     7201 ggagcgccgg cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt
     7261 gcgagagggc gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc
     7321 gccgccgcac cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa
     7381 tgggcgggga gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc
     7441 ggggctgtcc gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct
     7501 tctggcgtgt gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt
     7561 tcctacagct cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat
     7621 tcc
//
